1.Efficacy observation of acupuncture for dry eye syndrome of lung-yin deficiency pattern
Jing-Wen LIU ; Yao-Dong ZHAO ; Ling ZHU ; Cong HAN
Journal of Acupuncture and Tuina Science 2021;19(1):72-77
Objective: To observe the clinical efficacy of combining two needling manipulations, Er Long Xi Zhu (two dragons playing with a pearl) and Guo Yan Re (heat produced to reach the eyes), in treating dry eye syndrome (DES) of lung-yin deficiency pattern. Methods: Fifty-six eligible DES patients of lung-yin deficiency were randomized into an observation group and a control group, with 28 cases in each group. Same acupoints were selected in the two groups: Cuanzhu (BL 2), Sizhukong (TE 23), Taiyang (EX-HN 5) and Fengchi (GB 20) were chosen as the major points and Feishu (BL 13) and Chize (LU 5) as the adjuvant. Er Long Xi Zhu and Guo Yan Re needling manipulations were applied in the observation group while twirling reinforcing manipulation was used in the control group. Treatment was conducted once per day in both groups, for two sets of 15 consecutive days at a 2-day interval. Changes in the symptom score, tear break-up time (BUT) and tear production were observed afterwards, and the clinical efficacy was also compared between the two groups. Results: The total effective rate was 92.8% in the observation group, higher than 71.4% in the control group, and the between-group difference was statistically significant (P<0.05). After treatment, the symptom score, tear BUT and tear production showed significant improvements in both groups (all P<0.05); the symptom score, BUT and tear production in the observation group were significantly different from those in the control group (all P<0.05). Conclusion: Given the same acupoint selection, combining Er Long Xi Zhu and Guo Yan Re needling manipulations can produce more significant clinical efficacy than twirling reinforcing manipulation in treating DES of lung-yin deficiency pattern.
2.Effect of bicuculline on first spike latency of the neurons in the inferior colliculus of mice.
Journal of Southern Medical University 2009;29(5):918-921
OBJECTIVETo evaluate the effect of bicuculline on the first spike latency of the neurons in the inferior colliculus of mice and investigate the role of GABA inhibition in sound signal processing of the neurons.
METHODSIn vivo extracellular recording was performed on the inferior colliculus of 13 BALB/c mice (4 to 5 weeks old) to record the neuronal response to pure tones. Bicuculline, a GABA-A receptor antagonist, was applied to the neurons iontophoretically through one channel in the three-barrel glass-pipettes. The first spike latency and other response properties of the characteristic frequency were recorded for analysis.
RESULTSA total of 30 well-isolated single neurons were recorded. Increased spike counts characterized 96% of the neurons, with either increased (40%) or decreased (60%) latency of neuronal responses. Characteristic frequency alterations occurred in 50% of the neurons with increased spike latency, and the minimum threshold showed linear changes.
CONCLUSIONGABAergic inhibition may participate in the latency formation and increased frequency selectivity of mouse inferior colliculus neurons by lateral inhibition. The changes in the first spike latency can be indicative of the information integration in GABAergic neurons at the synaptic level.
Acoustic Stimulation ; psychology ; Action Potentials ; drug effects ; Animals ; Bicuculline ; pharmacology ; Female ; GABA Antagonists ; pharmacology ; Inferior Colliculi ; physiology ; Male ; Mice ; Mice, Inbred BALB C ; Neurons ; physiology ; gamma-Aminobutyric Acid ; physiology
3.Effects of p300/CBP on histone acetylation of Foxp3 gene in children with Kawasaki disease
Jiehua MEI ; Qin WANG ; Guobing WANG ; Pengqiang WEN ; Mingguo XU ; Gen TANG ; Dong CUI ; Cong LIU ; Dongli MA ; Chengrong LI
Chinese Journal of Microbiology and Immunology 2017;37(5):347-354
Objective To investigate the effects of p300/CBP on histone acetylation of Foxp3 gene and its roles in the immunological pathogenesis of Kawasaki disease (KD).Methods Forty-six children with KD and twenty-eight age-matched health children were consented to participate in this study.Co-immunoprecipitation and real-time PCR were performed to detect Foxp3-associated acetylation levels of histone H4 and binding abilities of p300, CBP, pSmad3 (phosphorylated mothers against decapentaplegic homolog 3) and NF-AT (nuclear factor of activated T cells) with Foxp3 gene in CD4+ T cells.The percentages of CD4+CD25high Foxp3+ cells (Treg) and the expression of Foxp3, CTLA4 (cytotoxic T-lymphocyte-associated protein 4), p300, CBP, TGF-βRⅡ (transforming growth factor β receptor Ⅱ) and pLAT1 at protein level were analyzed by flow cytometry.Quantitative real-time PCR was used to measure the expression of Foxp3, IL-10, TGF-β, TGF-βRⅠ, Egr-1 (early growth response protein 1), RARα (retinoic acid receptor α) and PLCγ1 (phospholipase C-γ1) in Treg cells at mRNA level.Plasma concentrations of TGF-β and retinol acid (RA) were measured by enzyme-linked immunosorbent assay.Results (1) The percentages of Treg cells, levels of Foxp3 and molecules associated with suppressive function of Treg cells (TGF-β, IL-10 and CTLA4), acetylation levels of histone H4 associated with promoter, conserved non-coding DNA sequence 1 (CNS1) and CNS2 of Foxp3 gene decreased remarkably during acute KD (P<0.05), but were restored after IVIG therapy (P<0.05).Meanwhile, all of the aforementioned items in KD patients with coronary artery lesions (KD-CAL+) were lower than those without coronary artery lesions (KD-CAL-) (P<0.05).No significant differences in histone H4 acetylation associated with CNS3 were found among different groups (P>0.05).(2) The levels of p300 and CBP in Treg cells and their binding abilities with Foxp3 gene were down-regulated significantly during acute KD (P<0.05), but were restored to some extent after IVIG treatment (P<0.05).The Foxp3-associated histone acetylation was positively correlated with the expression of p300 and CBP at mRNA level during acute KD (r=0.65, 0.42, P<0.05).Furthermore, the expression of p300 and CBP and their binding abilities with Foxp3 gene in KD-CAL+ group were lower than those in KD-CAL-group (P<0.05).(3) Compared with healthy subjects, plasma concentrations of TGF-β and RA and the expression of TGF-βRⅠ/Ⅱ/Egr-1, RARα and pLAT1/PLCγ1 were down-regulated during acute KD (P<0.05);the binding abilities of pSmad3 and NFAT with Foxp3 gene were reduced remarkably in patients with acute KD (P<0.05).All the items mentioned above were restored after IVIG treatment (P<0.05).Moreover, the ten items aforementioned in KD-CAL+ group were lower than those in KD-CAL-group (P<0.05).(4) Higher acetylation levels of histone H4 associated with promoter, CNS1 and CNS2, and enhanced binding abilities of p300 and CBP with Foxp3 gene were found in CD4+ T cells isolated from patients with acute KD after co-stimulation with TGF-β, RA and anti-CD3/CD28 antibodies as compared with those in CD4+ T cells without stimulation (P<0.05).However, no statistical difference in the acetylation level of histone H4 associated with CNS3 was found between the two groups (P>0.05).Conclusion Hypoacetylation of histone H4 associated with Foxp3 gene caused by insufficient expression of p300/CBP and their impaired binding abilities might be involved with immune dysfunction in KD.IVIG therapy regulates the expression of p300/CBP and their binding abilities with Foxp3 gene through up-regulating TGF-β signal.
4.Effects of SMYD3 and MLL5 on histone methylation of Foxp3 gene in children with Kawasaki disease
Jiehua MEI ; Qin WANG ; Guobing WANG ; Pengqiang WEN ; Mingguo XU ; Gen TANG ; Dong CUI ; Cong LIU ; Dongli MA ; Chengrong LI
Chinese Journal of Rheumatology 2017;21(8):518-523
Objective To investigate the effects of SMYD3 and MLL5 on histone methylation of Transcription factor forkhead box protein 3 (Foxp3) gene and its roles in the immunological pathogenesis of Kawasaki disease (KD). Methods Forty-two children with KD and 26 age-matched healthy children were consented to participate in this study. Co-Immunoprec-ipitation and real-time polymerase chain reaction (PCR) was performed to determine Foxp3-associated histone methylation levels of H3K4me3 and H3K27me3, and binding levels of SMYD3 and MLL5 with Foxp3 gene in CD4+T cells. The proportion of CD4+CD25high Foxp3+cells (Treg) and protein levels of Foxp3, cytotoxic T lymphocyte associated antigen-4 (CTLA4), TGF-βRⅡand pSmad3 were analyzed by flow cytometry. Quantitative real-time PCR was used to evaluate levels of Foxp3, interleukin (IL)-10, GITR, TGF-βRⅠand RARαmRNA in CD4+T cells. Plasma concentrations of TGF-βand retinol acid (RA) were measured by enzyme-linked Immunosorbent assay. Independent-samples t-test was used as the statistical method in this study. Results ① The proportion of Treg, expression levels of Foxp3 and molecules associated with suppressive function of Treg cells(IL-10, GITR and CTLA4), and histone methylation levels of H3K4me3 associating with promoter, conserved non-coding DNA sequence (CNS) 1 and CNS2 of Foxp3 gene decreased remarkably during acute KD [Promoter:(5.4±1.8)%vs (9.1±2.2)%;CNS1:(2.6±0.9)% vs (3.8±1.1)%; CNS2: (2.4±0.8)% vs (4.2±1.0)%; t=5.50, 6.02, 9.56, 7.92, 7.97, 4.76, 7.73, 5.01, 8.66; P<0.05], and restored after intravenous immunoglobulins (IVIG) therapy [Promoter: (7.2 ±2.1)% vs (5.4 ±1.8)%; CNS1:(3.6±1.4)% vs (2.6±0.9)%; CNS2: (3.6±1.4)% vs (2.4±0.8)%; t=5.56, 4.59, 7.01, 6.04, 5.89, 4.83, 4.45, 4.00, 5.12; P<0.05]. Meanwhile, the nine former items in KD patients with coronary artery lesions (KD-CAL+) were lower than those without coronary artery lesions (KD-CAL-) [Promoter: (4.11±1.45)% vs (6.16±1.93)%; CNS1:(1.99±0.87)%vs (2.96±1.10)%;CNS2: (1.75±0.63)%vs (2.72±1.16)%;t=6.28, 3.24, 4.56, 3.69, 3.38, 4.40, 3.65, 3.00, 3.51; P<0.05]. No significant difference of H3K4me3 associated with CNS3 and H3K27me3 were found among the groups (t=1.03, 0.91, 1.48 and 0.79, 0.82, 1.53; P>0.05). ② Binding levels of SMYD3 and MLL5 with Foxp3 gene in CD4+T cells were down-regulated significantly during acute KD (t=6.63, 6.15; P<0.05), and restored to some extent after IVIG treatment (t=5.36, 4.56; P<0.05). Positive correlations between binding levels of SMYD3 and MLL5 and expression level of Foxp3 mRNA were detected in patients with acute KD (r=0.62、0.45, P<0.05). Furthermore, Binding levels of SMYD3 and MLL5 with Foxp3 gene in KD-CAL+group were lower than those in KD-CAL- group (t=4.11, 4.31; P<0.05). ③ Compared with healthy controls, plasma concentration of TGF-β and RA, and expressions of TGF-βRⅡ, TGF-βRⅠ, pSmad3 and RARα were down-regulated during acute KD (t=11.54, 12.81, 7.43, 16.10, 8.25, 12.06; P<0.05), and elevated remarkably after IVIG treatment (t=8.40, 6.24, 5.94, 11.78, 6.27, 8.30; P<0.05). Simultaneously, all the items aforementioned in KD-CAL+ group were found to be lower than those in KD-CAL-group (t=3.58, 3.30, 3.82, 5.27, 4.71, 3.78; P<0.05). Conclusion Hypomethylation of H3K4me3 associated with Foxp3 gene caused by insufficient binding levels of SMYD3/MLL5 may be involved with immune dysfunction in Kawasaki disease.
5.Changes and significances of IL-17-associated histone methylation in the acute phase of Kawasaki dis-ease
Jiehua MEI ; Qin WANG ; Pengqiang WEN ; Mingguo XU ; Gen TANG ; Dong CUI ; Cong LIU ; Chengrong LI ; Guobing WANG
Chinese Journal of Microbiology and Immunology 2016;36(9):692-698
Objective To investigate the changes and significances of IL-17-associated histone methylation in the acute phase of Kawasaki disease (KD). Methods Forty-two children with KD and 28 age-matched healthy children were recruited in this study. Co-immunoprecipitation and real-time PCR were performed to detect the IL-17-associated histone methylation in CD4+ T cells. The percentages of CD4+IL-17+ T cells (Th17) and the expression of IL-17 and pSTAT3 at protein level were analyzed by flow cytome-try. Quantitative real-time PCR was used to measure the expression of IL-17, IL-6Rα, gp130, IL-23R, IL-23Rβ1, ETV5, SOCS1, SOCS3, TLR4, MyD88/TRIF, TNFR1 and RIP1 at mRNA level and the expres-sion of miR155 in CD4+ T cells. The levels of IL-6, IL-23 and TNF-αin plasma samples were measured by enzyme-linked immunosorbent assay. Results (1) The children with acute KD showed increased percenta-ges of Th17 cells and enhanced expression of IL-17 and H3K4me3, but inhibited expression of H3K27me3 [H3K4me3:(3.79±1. 45)% vs (1. 93±0. 31)%, H3K27me3: (54. 51±13. 60)% vs (73. 96± 22. 32)%;P<0. 05]. Moreover, the three former indexes in KD patients complicated with coronary artery lesions ( KD-CAL+) were higher than those in KD patients without coronary artery lesions ( KD-CAL-) , while the levels of H3K27me3 in KD-CAL+ group were lower than those in KD-CAL- group [ H3K4me3:(5. 11±1. 68)% vs (2. 98±0. 99)%, H3K27me3:(45. 02±14. 83)% vs (60. 35±12. 51)%;P<0. 05]. A positive correlation was observed between the ratio of H3K4me3/H3K27me3 and IL-17 at transcriptional level in patients with acute KD (r=0. 69, P<0. 05). Treatment with intravenous immunoglobulin (IVIG) restored Th17 cells, expression of IL-17 and methylation levels of H3K4me3 and H3K27me3 to normal levels [H3K4me3:(2. 44 ± 0. 77)% vs (3. 79 ± 1. 45)%, H3K27me3: (66. 52 ± 15. 73)% vs (54. 51 ± 13. 60)%;P<0. 05]. (2) The expressions of pSTAT3, ETV5 and miR155 increased significantly in pa-tients with acute KD, while the expressions of negative regulators of pSTAT3 ( SOCS1 and SOCS3 ) were down-regulated. The expressions of pSTAT3, ETV5 and miR155 in KD-CAL+ group were higher than those in KD-CAL- group (P<0. 05), while the levels of SOCS1 and SOCS3 in KD-CAL+ group were lower than those in KD-CAL- group (P<0. 05). IVIG therapy restored the indexes mentioned above to some extent (P<0. 05). (3) Compared with the healthy subjects, the levels of inflammatory cytokines (IL-6, IL-23 and TNF-α) in plasma and the expressions of surface receptors (TLR4, IL-6Rα/gp130, IL-23R/IL-23Rβ1 and TNFR1) and its downstream adaptors (MyD88, TRIF, RIP1) in CD4+T cells were up-regulated in patients with acute KD (P<0. 05), but were down-regulated significantly after IVIG treatment (P<0. 05). Moreo-ver, all of the indexes mentioned above in KD-CAL+ group were found to be higher than those in KD-CAL-group (P<0. 05). Conclusion Aberrant patterns of IL-17-associated histone methylation might be related to the immune dysfunction in patients with KD.
6.Calcium transient of CA1 pyramidal neurons induced by potassium blocker 4-aminopyridine in acute hippocampal slices
Tao SU ; Wen-Dong CONG ; Wei-Ping LIAO
Chinese Journal of Neuromedicine 2011;10(5):467-470
Objective To investigate the calcium transient of CA1 pyramidal neurons induced by potassium blocker 4-aminopyridine (4-AP) in acute hippocampal slices to explore the relation between potassium channel function and calcium transient, and their mechanism. Methods Fluorescent probe was employed to mark the hippocampai neurons in acute brain slices of rats; confocal microscopy was used to perform calcium imaging to observe the influences of different concentrations of 4-AP and perfusate with/without calcium on calcium transient of CA1 pyramidal neurons. Results The response of [Ca2+]I to lower concentration of 4-AP (<15 mmol/L) was in a dose-dependent manner (r2=0.910, P=0.000); the higher the concentration of 4-AP (20-80 mmol/L), the lower the peak level of calcium transient. The latency and amplitude of calcium transient induced by 4-AP were obviously reduced when the extracellular condition was switched to an absence of calcium, which was significantly different as compared with that with calcium (P<0.05). Conclusion Blockade of potassium channels with 4-AP can increase [Ca2+]I in the hippocampal pyramidal neurons of acute slices. The increase of [Ca2+]1 to 4-AP could be ascribe to calcium release from intracellular stores and calcium influx from extracellular matrix.
7.Uniformity of Coptidis Rhizoma decoction pieces dispensing based on effective constituent equivalence.
Qin DONG ; Jia-bo WANG ; Ding-kun ZHANG ; Wu-wen FENG ; Cong-en ZHANG ; Ming NIU ; Xiao-ping DONG ; Xiao-he XIAO
China Journal of Chinese Materia Medica 2015;40(20):3981-3986
Traditional Chinese medicine (TCM) dispensing is the final step of TCM used for clinical treatment, the stability of TCM dispensing is the guarantee of good clinical effect. Establishment of effect-constituent equivalence for Chinese herbal pieces based on clinical efficacy, can not only guarantee the stability of TCM dispensing, but also relate to the precision of clinical effect. This study chose Coptidis Rhizoma as the model, established effect-constituent equivalence of Coptidis Rhizoma, based on the effect-constituent index already established by our research group, and taking into consideration of homogeneity of clinical dosage and compliance of decoction, the uniformity of dispensing for different specification of Coptidis Rhizoma decoction pieces was studied. This research model was then applied to guide the specification-optimization of Coptidis Rhizoma and its clinical dispensing. The result indicated, effective constituent equivalence could reflect the fluctuation of specification, dosage and decoction to the fluctuation of efficacy; Optimized Coptidis Rhizoma decoction pieces had the characteristic of high homogeneity as for clinical dispensing, good compliance as for decoction, and high effective constituent equivalence. In conclusion, effective constituent equivalence could improve relevance of methods of TCM dispensing control to clinical effect. Preparated Superior-standard Decoction Pieces based on effective constituent equivalence was featured by good quality and a good practice of adjustable dosage, which could promote the development of TCM decoction pieces toward precision medicine.
Chemistry, Pharmaceutical
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methods
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Coptis
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chemistry
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Drugs, Chinese Herbal
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chemistry
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Quality Control
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Rhizome
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chemistry
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Therapeutic Equivalency
8.Evaluation of immunohistochemical markers for differential diagnosis of hepatocellular carcinoma from intrahepatic cholangiocarcinoma.
Hui DONG ; Wen-Ling CONG ; Zhong-Zheng ZHU ; Bin WANG ; Zhi-Hong XIAN ; Hua YU
Chinese Journal of Oncology 2008;30(9):702-705
OBJECTIVETo evaluate the significance of a panel of immunohistochemical markers for distinguishing hepatocellular carcinoma (HCC) from intrahepatic cholangiocarcinoma (ICC).
METHODSTen markers including hepatocyte paraffin 1 (Hep Par 1), polyclonal carcinoembryonic antigen (pCEA), CD34, CD10, CD105, multidrug resistance-associated protein-3 (MRP-3), cyclooxygenase-2 (COX-2), mucinous glycoprotein-1 (MUC-1), aquaporin-1 (AQP-1) and CK19 were immunohistochemically stained in the samples from 90 surgically resected HCC and 80 ICC, respectively,and the positive rate of their expression were compared statistically.
RESULTSThe positive expression rates of Hep Par 1, pCEA, CD34, CD10, CD105, MRP-3, COX-2 were 85.6%, 82.2%, 87.8%, 18.9%, 8.9%, 11.1% and 48.9%, respectively, in HCC. While the positive expression rates of MUC-1, AQP-1 and CK19 were 73.8%, 65% and 92.5%, respectively, in ICC.
CONCLUSIONBased on our results, Hep Par 1 and CD34 can be used as the first line markers, and pCEA and COX-2 as the second line makers, for differential diagnosis of hepatocellular carcinoma from intrahepatic cholangiocarcinoma. While MUC-1 and CK19 can be used as the first line markers and AQP-1 as the second one for the differential diagnosis of intrahepatic cholangiocarcinoma from hepatocellular carcinoma.
Bile Duct Neoplasms ; chemistry ; diagnosis ; Bile Ducts, Intrahepatic ; chemistry ; Biomarkers, Tumor ; analysis ; Carcinoma, Hepatocellular ; chemistry ; diagnosis ; Cholangiocarcinoma ; chemistry ; diagnosis ; Diagnosis, Differential ; Female ; Hepatocytes ; chemistry ; pathology ; Humans ; Immunohistochemistry ; Liver Neoplasms ; chemistry ; diagnosis ; Male ; Middle Aged
9.A novel mutation of the KIT gene in a Chinese family with piebaldism.
Guang-Dong WEN ; Cheng ZHOU ; Cong YU ; Juan DU ; Qian-Xi XU ; Zheng-Yi LIU ; Jian-Zhong ZHANG
Chinese Medical Journal 2013;126(12):2325-2328
BACKGROUNDHuman piebaldism is a rare autosomal dominant condition characterized by congenital white forelock and depigmented patches of skin, typically on the forehead, anterior trunk and extremities. Mutations in the KIT gene have been proposed to be responsible for the underlying changes in this disorder. The aim of this study was to identify gene mutation in a Chinese family with piebaldism.
METHODSA Chinese family with piebaldism presenting with white forelock and large depigmented skin macules on the abdomen, arms and legs was collected. DNA was isolated from peripheral blood of the family members. The encoding exons with flanking intron regions of the KIT gene were analyzed by polymerase chain reactions (PCR) and direct DNA sequencing. Besides, DNA extracted from 100 ethnically matched population individuals was as controls.
RESULTSA heterozygous missense mutation c.2590T > C was identified in the patients of the family. This mutation converted a serine residue to proline (p.Ser864Pro). The mutation was not found in their unaffected family members or normal controls.
CONCLUSIONA novel missense mutation c.2590 T > C was found and it might play a significant role in the piebaldism phenotype in the family.
Child ; Humans ; Male ; Mutation, Missense ; Piebaldism ; genetics ; Proto-Oncogene Proteins c-kit ; genetics ; physiology
10.Partial AZfc region deletions of the Y chromosome in spermatogenic dysfunction patients.
Xue-Wen CONG ; Xiang-Dong TU ; Ai-Zhen YAN ; Jian ZENG
National Journal of Andrology 2010;16(7):594-598
OBJECTIVETo investigate the influence of partial deletions in the AZFc region of the Y chromosome on spermatogenesis.
METHODSWe selected 9 sequence tagged sites (sY1258, sY1291, sY254, sY255, sY1201, sY1206, sY1161, sY1197 and sY1191) in the AZFc region of the Y chromosome, with ZFX/ZFY and SRY (sY14) as the interior control. We amplified by multiplex PCR the DNA of 160 patients with azoospermia or severe oligozoospermia that showed no microdeletion of the Y chromosome (the case group) and another 76 males with normal fertility (the control group). For the individuals suspected of DAZ gene deletion, we detected the single nucleotide polymorphism sites (SNPs) in the four copies of the DAZ gene by single nucleotide variation (SNV) analysis to determine the types of DAZ copy deletion.
RESULTSIn the case group, there were 10 cases of gr/gr (sY1291) deletion (6.3%), 14 b2/b3 (sY1191) deletion (8.8%), 1 sY1291,sY1197 deletion (0.6%), 1 b1/b2 deletion (0.6%) and 1 b1/b3 deletion (0.6%), while in the control group, there were 4 cases of gr/gr deletion (5.3%) and 4 b2/b3 deletion (5.3%). SNV analysis showed DAZ1/DAZ2 deletion in all those with gr/gr and those with b1/b3 deletion, DAZ3/DAZ4 deletion in those with b2/b3 deletion, and DAZ-SNV sY587 deletion in 1 case of sY1291, sY1197 deletion, but no DAZ deletion was found in 1 case of b1/b2 deletion.
CONCLUSIONB2/b3 (sY1191) and gr/gr (sY1291) deletions are genomic polymorphisms and quite common in the normal Chinese population; while b1/b2, b1/b3, and sY1291, sY1197 deletions may be high risk factors of dyszoospermia.
Case-Control Studies ; Chromosomes, Human, Y ; Humans ; Male ; Oligospermia ; genetics ; Sequence Deletion ; Spermatogenesis